Abstract
Covalent ligands are a versatile class of chemical probes and drugs that can target noncanonical sites on proteins and display differentiated pharmacodynamic properties. Chemical proteomic methods have been introduced that leverage electrophilic fragments to globally profile the covalent ligandability of nucleophilic residues, such as cysteine and lysine, in native biological systems. Further optimization of these initial ligandability events without resorting to the time-consuming process of individualized protein purification and functional assay development, however, presents a persistent technical challenge. Here, we show that broadly reactive electrophilic fragments, or “scouts”, can be converted into site-specific target engagement probes for screening small molecules against a wide array of proteins in convenient gel- and ELISA-based assay formats. We use these assays to expediently optimize a weak potency fragment hit into a sub-μM inhibitor that selectively engages an active-site cysteine in the retinaldehyde reductase AKR1B10. Our findings provide a road map to optimize covalent fragments into more advanced chemical probes without requiring protein purification or structural analysis.
Highlights
Covalent ligands are a versatile class of chemical probes and drugs that can target noncanonical sites on proteins and display differentiated pharmacodynamic properties
Chemical proteomic strategies have emerged that enable the discovery of ligands for proteins on a global scale directly in native biological systems.[7−10] A subset of these approaches focuses on covalent ligand discovery, where electrophilic compounds are screened for reactivity against thousands of nucleophilic residues in the proteome.[11,12]
Covalent ligands are attractive starting points for chemical probe and drug development for several reasons, including the potential to achieve improved potency for shallow binding pockets and increased residency time leading to more durable pharmacological action.[13−15] Covalent chemical probes and drugs have been developed against diverse proteins, including hydrolases,[16,17] kinases,[18−20] nuclear export proteins,[21] and oncogenic GTPases.[22,23]
Summary
Covalent ligands are a versatile class of chemical probes and drugs that can target noncanonical sites on proteins and display differentiated pharmacodynamic properties.
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